3T3-Ll fibroblasts differentiate in culture to develop phenotypic characteristics of adipocytes. Differentiation can be induced by incubation of confluent fibroblasts for 72 hrs with insulin, IBMX and dexamethasone. Soluble cAMP PDE activity in undifferentiated fibroblasts as well as adipocytes is very sensitive to inhibition by Ro 20-1724, but not cilostamide, whereas adipocyte particulate cAMP PDE is very sensitive to inhibition by cilostamide, but not Ro 20-1724. IBMX readily inhibits both soluble and particulate cAMP PDE activities. Ro 20-1724, but not cilostamide, can substitute for IBMX in inducing differentiation. These and other experiments suggest an important role for soluble PDE in the regulation of differentiation. In adipocytes, cilostamide enhanced basal and isoproterenol-stimulated lipolysis to a greater extent than Ro 20-1724, but had a lesser effect on cAMP content than RO 20-1724. The antilipolytic effect of insulin was inhibited by cilostamide but not Ro 20-1724. These findings suggest that particulate cAMP PDE is important in metabolism of cAMP involved in regulation of lipolysis, as well as in the antilipolytic action of insulin. Incubation of adipocytes with isoproterenol or the antilipolytic agents insulin and PIA for 6-10 min resulted in activation of the particulate cAMP PDE. PIA also inhibited isoproterenol-stimulated adenylate cyclase activity. The effects of insulin and PIA, but not isoproterenol, on PDE activation were inhibited by pertussis toxin, suggesting a role for guanyl nucleotide binding proteins in PDE regulation and/or insulin action. Insulin did not apparently increase hydrolysis of phosphatidyl inositol or activate protein kinase C. Phorbol ester did induce "translocation" of protein kinase C but did not alter insulin's effects on PDE, lipolysis, or glucose transport.